Transcriptional profilling of hESC with and without 17q gain upon induction of replicative stress through HU treatment [targeted exome sequencing]

Human embryonic stem cells (hESC) and cancer cells rapidly divide with a short G1/S-phase causing increased replicative stress (RS). Since both in vitro cultured hESCs and most high-risk neuroblastomas have large chromosome 17q gains (17q+), we hypothesize that this may provide a "RS-resistance phenotype". We co-cultured parental cells and a derived hESC line with 17q+ under normal growth conditions and under RS. We could show a proliferative ad-vantage of hESC with 13q+17q+ over wild type by measurement of the cumulative growth and molecular analysis for chromosomal copy number analysis. To monitor effects of 17q+ on RS-resistance, cell cycle and transcriptome analysis were performed. In conclusion, we show that extra chromosomal aberrations, such as 17q+, provide proliferative advantage to hESC under RS and suggest that this phenomenon explains the high incidence of 17q+ in in vitro cultured hESC lines. Samples were collected at each splitting moments of hESC with and without 17q gain, and of the 1:1 mixed culture; under normal growth conditions (PBS) or conditions of replicative stress (150 µM HU), for 2 biological replicates

人类胚胎干细胞（human embryonic stem cells, hESC）与癌细胞均以快速分裂为特征，且G1/S期较短，由此引发复制应激（replicative stress, RS）水平升高。鉴于体外培养的人类胚胎干细胞以及多数高危神经母细胞瘤均存在17号染色体长臂q区大片段扩增（17q+），我们推测该特征可能赋予细胞“复制应激抵抗表型”。我们在正常培养条件与复制应激条件下，分别共培养亲本细胞与一株携带17q+的人类胚胎干细胞衍生系。通过累积生长检测与染色体拷贝数分子分析，我们证实携带13q+17q+的人类胚胎干细胞较野生型细胞具有增殖优势。为探究17q+对复制应激抵抗能力的影响，本研究开展了细胞周期与转录组分析。综上，本研究证实，诸如17q+这类染色体畸变可在复制应激条件下赋予人类胚胎干细胞增殖优势，且该现象可解释体外培养人类胚胎干细胞系中17q+的高发生率。本研究设置2次生物学重复，分别收集携带与不携带17q扩增的人类胚胎干细胞、以及1:1混合共培养细胞在每次传代时的样本；培养条件包括正常生长条件（磷酸盐缓冲液，PBS）与复制应激条件（150 μM羟基脲，HU）。